ABSTRACT
Tuberculosis is a major global cause of both mortality and financial burden mainly in low and middle-income countries. Given the significant and ongoing rise of drug-resistant strains of Mycobacterium tuberculosis within the clinical setting, there is an urgent need for the development of new, safe and effective treatments. Here the development of a drug-like series based on a fused dihydropyrrolidino-pyrimidine scaffold is described. The series has been developed against M. tuberculosis lysyl-tRNA synthetase (LysRS) and cellular studies support this mechanism of action. DDD02049209, the lead compound, is efficacious in mouse models of acute and chronic tuberculosis and has suitable physicochemical, pharmacokinetic properties and an in vitro safety profile that supports further development. Importantly, preliminary analysis using clinical resistant strains shows no pre-existing clinical resistance towards this scaffold.
Subject(s)
Lysine-tRNA Ligase , Mycobacterium tuberculosis , Tuberculosis , Animals , Lysine-tRNA Ligase/chemistry , Lysine-tRNA Ligase/genetics , Lysine-tRNA Ligase/pharmacology , Mice , Mycobacterium tuberculosis/genetics , Tuberculosis/drug therapyABSTRACT
Wild-type Escherichia coli K-12 was grown in minimal medium alone or with the addition of 20 mM L-alanine or 3 mM glycyl-L-leucine. A lysyl-tRNA synthetase mutant strain was grown in minimal medium containing 20mM L-alanine. The lysyl-tRNA synthetase from these strains was purified to 70-90% of homogeneity. Kinetic studies comparing the effect of thermal and urea inactivation on these different lysyl-tRNA synthetase preparations and measurement of the Michaelis constant for lysine and transfer RNA indicated that growth of Escherichia coli in the presence of alanine and glycyl-L-leucine induces an alteration in the properties of the synthetase. Measurement of the apparent Km for ATP at pH 7.25 indicates lysyl-tRNA synthetase has two two binding sites for this substrate, and further studies indicated a dependence of the apparent Km for lysine on the ATP concentration.